Opinion TRENDS in Plant Science Vol.11 No.1 January 2006

Abiotic stress, the field environment and stress combination

Ron Mittler

Department of Biochemistry and Molecular Biology, University of Nevada, Mail Stop 200, Reno, NV 89557, USA

Farmers and breeders have long known that often it is enhanced tolerance to a particular biotic or abiotic stress the simultaneous occurrence of several abiotic stresses, condition failed to show enhanced tolerance when tested rather than a particular stress condition, that is most in the field [12–14]. A focus on molecular, physiological lethal to crops. Surprisingly, the co-occurrence of and metabolic aspects of stress combination is needed to different stresses is rarely addressed by molecular bridge this gap and to facilitate the development of biologists that study plant acclimation. Recent studies crops and plants with enhanced tolerance to field have revealed that the response of plants to a stress conditions. combination of two different abiotic stresses is unique and cannot be directly extrapolated from the response of Tailoring a response to a particular stress situation plants to each of the different stresses applied individu- Plant acclimation to a particular abiotic stress condition ally. Tolerance to a combination of different stress requires a specific response that is tailored to the precise conditions, particularly those that mimic the field environmental conditions the plant encounters. Thus, environment, should be the focus of future research molecular, biochemical and physiological processes set in programs aimed at developing transgenic crops and motion by a specific stress condition might differ from plants with enhanced tolerance to naturally occurring those activated by a slightly different composition of environmental conditions. environmental parameters. Illustrating this point are transcriptome profiling studies of plants subjected to Abiotic stress research: a reality check different abiotic stress conditions: each different stress Abiotic stress conditions cause extensive losses to condition tested prompted a somewhat unique response, agricultural production worldwide [1,2]. Individually, and little overlap in transcript expression could be found stress conditions such as drought, salinity or heat have between the responses of plants to abiotic stress been the subject of intense research [2,3]. However, in the conditions such as heat, drought, cold, salt, high light field, crops and other plants are routinely subjected to a or mechanical stress [10,15–18].Althoughreactive combination of different abiotic stresses [4–7]. In drought- oxygen species (ROS) are associated with many different stricken areas, for example, many crops encounter a biotic or abiotic stress conditions, different genes of the combination of drought and other stresses, such as heat or ROS gene network of Arabidopsis were found to respond salinity [4,5]. Recent studies have revealed that the differently to different stress treatments [19]. These molecular and metabolic response of plants to a combi- findings suggest that each abiotic stress condition nation of drought and heat is unique and cannot be requires a unique acclimation response, tailored to the directly extrapolated from the response of plants to each of specific needs of the plant, and that a combination of two these different stresses applied individually [8–11]. or more different stresses might require a response that Studies of simultaneous stress exposure in different plants is also unique. are well documented in various agronomic and horticul- In addition to the basic differences that exist between ture journals. In addition, tolerance to a combination of the acclimation responses of plants to different abiotic two different abiotic stresses is a well-known breeding stress conditions [10,15–18], when combined, different target in corn and other crops [5–7]. Nevertheless, little is stresses might require conflicting or antagonistic known about the molecular mechanisms underlying the responses. During heat stress, for example, plants open acclimation of plants to a combination of two different their stomata to cool their leaves by transpiration. stresses [10]. Because the majority of abiotic stress studies However, if heat stress is combined with drought, plants performed under controlled conditions in the laboratory do would not be able to open their stomata and their leaf not reflect the actual conditions that occur in the field, a temperature would be higher [9]. Salinity or heavy considerable gap might exist between the knowledge metal stress might pose a similar problem to plants gained by these studies and the knowledge required to when combined with heat stress because enhanced develop plants and crops with enhanced tolerance to field transpiration could result in enhanced uptake of salt conditions. This gap might explain why some of the or heavy metals. Cold stress or drought, combined with transgenic plants developed in the laboratory with high light conditions, result in enhanced production of

Corresponding author: Mittler, R. ([email protected]). ROS by the photosynthetic apparatus because these Available online 15 November 2005 conditions limit the availability of CO2 for the dark www.sciencedirect.com 1360-1385/$ - see front matter Q 2005 Elsevier Ltd. All rights reserved. doi:10.1016/j.tplants.2005.11.002 16 Opinion TRENDS in Plant Science Vol.11 No.1 January 2006 reaction, leaving oxygen as one of the main reductive Case study: drought and heat stress products of photosynthesis [20]. Another example of Drought and heat stress represent an excellent example of antagonism between different abiotic stresses is drought two different abiotic stress conditions that occur in the and heavy metal stress, which exaggerate the effects of field simultaneously [5–7]. Several studies have examined each other [21]. Because energy and resources are the effects of a combination of drought and heat stress on required for plant acclimation to abiotic stress conditions the growth and productivity of maize, barley, sorghum and (e.g. for the synthesis of heat shock, or late embryogen- different grasses. It was found that a combination of esis abundant proteins), nutrient deprivation could pose drought and heat stress had a significantly greater a serious problem to plants attempting to cope with detrimental effect on the growth and productivity of heat, cold or drought stress. Likewise, limited avail- these plants and crops compared with each of the different ability of key elements such as iron, copper, zinc or stresses applied individually [5–7,23–27]. A sum of all manganese, which are required for the function of major US weather disasters between 1980 and 2004 different defense enzymes, such as superoxide dismutase (excluding hurricanes, tornadoes and wildfires; or ascorbate peroxidase, could result in an enhanced Figure 1a) demonstrates the extent of the damage caused oxidative stress in plants subjected to different abiotic by a combination of drought and heat stress [compare the stresses [22]. The acclimation of plants to a combination damage caused by drought to that caused by drought of different abiotic stresses would, therefore, require an combined with a heat wave in Figure 1a, and compare the appropriate response customized to each of the individ- maps showing vegetation health in Figure 1b: August ual stress conditions involved, as well as tailored to the 1997 (no drought, no heat wave), August 2000 (drought need to compensate or adjust for some of the antagon- combined with a heat wave) and August 2002 (drought istic aspects of the stress combination. without a heat wave)]. The vast agricultural areas

(a) (b) Percentage area dry 60 2 3 40 140 1 120 1 20

100 0 80 60

60 40 2

Billion US dollars 40 20 20

Percentage wet area wet Percentage 0 0 1996 1998 2000 2002 2004 Drought and Drought Freeze Flooding Year heat wave 1. August 1997 - No drought, no heat wave. 2. August 2000 - Drought and heat wave. 3 3. August 2002 - Drought, no heat wave.

(c) (d) (e)

Figure 1. The effects of a combination of drought and heat stress on US agriculture. (a) Total of all US weather disasters costing US$1 billion or more between 1980 and 2004 (excluding hurricanes, tornadoes and wildfires). Total damage was normalized to the 2002 US dollar value (http://www.ncdc.noaa.gov/oa/reports/billionz.html). (b) Vegetation health maps for three different times (end of August in 1997, 2000 and 2002) showing the effect of the combination of drought and heat (August 2000) on plant health. Left panel: percentage of area dry or wet between 1996 and 2004 in the USA; during 2000 and 2002 drought conditions were enhanced. Right panels: vegetation health maps corresponding to the times indicated by arrows in the left panel. Time point 1 (August 1997): no drought stress, no heat wave. Time point 2 (August 2000): a combination of drought and heat wave. Time point 3 (August 2002): drought no heat wave. (c) Temperature index map, (d) long-term drought – Palmer map, and (e) a satellite image of vegetation health for August 2000, a summer that included a drought and a heat wave causing damage of more than US$4.2 billion to US agriculture. Maps, statistical data and satellite images were obtained from the National Climatic Data Center (http://www.ncdc.noaa.gov). Data presented in Figure 1 should not be interpreted as scientific evidence for the effects of stress combination. Please see text and references below for controlled scientific experiments documenting the effects of stress combination on plant and crop performance. www.sciencedirect.com Opinion TRENDS in Plant Science Vol.11 No.1 January 2006 17 potentially affected by a combination of drought and heat stress can be extrapolated from weather maps and satellite images of the USA during August 2000, a (a) Transcripts (b) Metabolites month in which the co-occurrence of drought and heat Drought Heat Drought Heat stress caused damage costing more than US$4.2 billion (1571) (540) (23) (18) (Figure 1c). The data shown in Figure 1 support the findings described above [5–7,23–27], and vividly demon- 765 208 10 3 strate the impact that drought and heat stress have on 77 5 agriculture when combined. 729 255 8 10 Physiological characterization of plants subjected to drought, heat stress or a combination of drought and heat 772 5 stress reveals that the stress combination has several unique aspects, combining high respiration with low Drought + Heat Drought + Heat (1833) photosynthesis, closed stomata and high leaf temperature (28) (Figure 2) [9]. Starch breakdown coupled with energy TRENDS in Plant Science production in the mitochondria might, therefore, play a Figure 3. Unique molecular characteristics of drought and heat stress combination. key role in plant metabolism during a combination of Venn diagrams showing the overlap between (a) transcripts and (b) metabolites drought and heat stress [9,10]. Transcriptome profiling (enhanced or suppressed) during drought or heat stress, or a combination of studies of plants subjected to drought, heat stress or a drought and heat stress. The total number of transcripts or metabolites is indicated in parentheses. Data was obtained from [10]. combination of drought and heat stress support the physiological analysis of plants (Figure 2) [9,10] and suggest that the stress combination requires a unique are different and only a small overlap in transcript acclimation response involving O770 transcripts that are expression has been found between these responses. not altered by drought or heat stress (Figure 3) [10]. Overall, the example of drought and heat combination Similar changes in metabolite accumulation were also underlines the potential severity of a stress combination, found, with several unique metabolites, mainly sugars, as well as its unique physiological, molecular and accumulating specifically during the stress combination biochemical aspects. The results presented in Figures 1– (Figure 3) [10]. By contrast, the level of proline, thought to 3 strongly argue for a need to develop dedicated research be important for plant protection during drought stress programs aimed at enhancing the tolerance of plants and [2,3], is strongly suppressed during a combination of crops to a combination of different abiotic stresses. drought and heat stress [10]. The profiling experiments summarized in Figure 3 further illustrate that the Regulatory aspects of stress combination: a key to acclimation responses of plants to heat or drought stress enhancing tolerance? Enhancing plant tolerance to biotic or abiotic stress conditions by activating a stress-response signal transduc- tion pathway in transgenic plants is a powerful and 250 promising approach [3,28–30]. It is logical to assume that the simultaneous exposure of a plant to different abiotic stress conditions will result in the co-activation of 200 different stress-response pathways. These might have a synergisticorantagonisticeffectoneachother.In addition, dedicated pathways specific for the particular 150 stress combination might be activated [10]. Several examples of synergistic or antagonistic relationships between different pathways exist. Heat stress was found 100

% of Control to silence the UV-B response of parsley [31], whereas ozone induces the UV-B and/or pathogen responses of some other plants [32]. The phenomenon of cross-hard- 50 ening, which reflects some of the synergistic relationships among different stresses, has been reviewed in several papers (e.g. [33]). 0 Control Heat Drought Drought + Heat Cross-talk between co-activated pathways is likely to be mediated at different levels. These could include inte- Photosynthesis Stomatal conductance gration between different networks of transcription Respiration Leaf temperature factors and mitogen-activated protein kinase (MAPK)

TRENDS in Plant Science cascades [34,35], cross-talk mediated by different stress hormones such as ethylene, jasmonic acid and abscisic Figure 2. Unique physiological characteristics of drought and heat stress acid [36], cross-talk mediated by calcium and/or ROS combination. A combination of drought and heat stress is shown to be different from drought or heat stress by having a unique combination of physiological signaling [19,33], and cross-talk between different parameters. Data was obtained from [9,10]. receptors and signaling complexes [37]. Although evidence www.sciencedirect.com 18 Opinion TRENDS in Plant Science Vol.11 No.1 January 2006 exists for stress-mediated cross-talk at the level of MAPKs and hormone signaling [34–36], much remains to be studied, particularly if we wish to use specific signal

transduction components as a molecular leverage to Drought Salinity Heat Chilling Freezing Ozone Pathogen UV Nutrient enhance the tolerance of plants and crops to a combination Drought of different stresses. Ethylene was recently shown to play a central role in the response of Arabidopsis to a Salinity combination of heat and osmotic stress, and expression of the transcriptional co-activator MBF1c in Arabidopsis Heat was found to enhance the tolerance of transgenic plants to Chilling heat and osmotic stress by partially activating or perturbing the ethylene-response signal transduction Freezing pathway [11]. Ozone

Summary and conclusions: the ‘stress matrix’ Pathogen Theextentofdamagecausedtoagriculturebya UV combination of two different stresses (Figure 1) under- scores the need to develop crops and plants with enhanced Nutrient tolerance to a combination of different abiotic stresses. Drawing upon the limited physiological, molecular and metabolic studies performed with plants that were Potential negative interaction Unknown mode of interaction simultaneously subjected to two different abiotic stresses (Figures 2–3) [5–11,23–27], it is not sufficient to study Potential positive interaction No interaction each of the individual stresses separately. The stress combination should be regarded as a new state of abiotic TRENDS in Plant Science stress in plants that requires a new defense or acclimation Figure 4. Agriculturally important stress combinations (‘The Stress Matrix’). response. It should be studied in the laboratory or the field Different combinations of biotic and abiotic stresses are presented in the form of a matrix to demonstrate potential interactions that can have important implications by simultaneously exposing plants to different abiotic for agriculture. Different interactions are color coded to indicate potential negative stresses [10]. In addition, transgenic plants with enhanced [i.e. enhanced damage or lethality owing to the stress combination (purple)] or tolerance to biotic or abiotic stress conditions should be potential positive [i.e. cross-protection owing to the stress combination (green)] effects of the stress combination on plant health. However, the potential effects of tested for their tolerance to a combination of different stress combination could vary depending on the relative level of each of the stresses before they are introduced into field trials. different stresses combined (e.g. acute versus low) and the type of plant or We face many challenges in our attempts to develop pathogen involved. Data to generate the matrix was obtained from [4–10,21–23,25– 27,31–33,35,36,38–45]. transgenic plants with enhanced tolerance to a stress combination. Tolerance to a combination of different stresses is likely to be a complex trait involving multiple the biotic–abiotic axis, most of the different abiotic stress pathways and cross-talk between different sensors and combinations presented in Figure 4 have received almost signal transduction pathways. Mapping genes essential no attention. The experience of farmers and breeders for tolerance to a combination of abiotic stresses could, should be used as a valuable guide and resource to therefore, be costly and pose a technical challenge molecular biologists trying to address a specific stress requiring multiple controls. In addition, resistance to a combination that is pertinent to their crop of interest or combination of different stresses could be genetically region. The data presented in, for example, Figure 1, linked to suppressed growth or yield of plants. However, combined with different reports available from websites some stress combinations have the advantage of enhan- such as http://www.usda.gov/wps/portal/usdahome indi- cing lethality [25]. Genetic screens of mutant or inbred cate that major US crops, including corn and soybean, are lines could, therefore, be turned into selection for particularly vulnerable to a combination of drought and survivors with enhanced tolerance, and identified genes heat stress. By contrast, reports from northern countries could be tested in transgenic plants. such as Sweden or Canada identify a combination of cold What stress combinations should we study? Figure 4 stress and high light as having a major rate-limiting affect summarizes many of the stress combinations that could on agriculture. have a significant impact on agricultural production (The Perhaps the most important guideline for studying ‘Stress Matrix’). Perhaps the most studied interactions abiotic stress combination is to address it as if it is a new presented in Figure 4 are those between different abiotic state of abiotic stress in plants and not simply the sum of stresses and pests or pathogens (i.e. biotic stress). In some two different stresses. To develop transgenic crops with instances it has been reported that a particular abiotic enhanced tolerance to field conditions, researchers need to stress condition, such as ozone stress, enhances the widen their area of study to include stress combination. tolerance of plants to pathogen attack [38,39]. However, in most cases, prolonged exposure of plants to abiotic Acknowledgements stress conditions, such as drought or nutrient deprivation, Research in my laboratory is supported by funding from The National results in the weakening of plant defenses and enhanced Science Foundation (NSF-0431327; NSF-0420033) and The Nevada susceptibility to pests or pathogens [32,40]. In contrast to Agricultural Experimental Station (Publication number 03055517). www.sciencedirect.com Opinion TRENDS in Plant Science Vol.11 No.1 January 2006 19

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